CRISPR/Cas9 Targeting of Aldehyde Dehydrogenase 1A1 Reveals Heterogeneous Roles in Radiation Response and Redox Stress Across Clonal Lines in Triple-Negative Breast Cancer
Abstract
1. Introduction
2. Results
2.1. Effect of CRISPR/Cas9 Knockout (KO) of ALDH1A1 SUM159 Bulk Cells
2.2. Clonal Cell Lines Derived from Bulk Knockout Cells Have Decreased ALDEFLUORTM Activity
2.3. Clonal Differences in the Effect of ALDH1A1 Knockout on Cellular Metabolic Activity and Colony Formation
2.4. Clonal Differences in the Effect of ALDH1A1 Knockout on Viability, Apoptosis, and Reactive Oxygen Species
2.5. Radiation Sensitivity in ALDH1A1 Knockout Clones
3. Discussion
4. Materials and Methods
4.1. Cell Culturing
4.2. CRISPR/Cas9 Targeting
4.3. Western Blot Analysis
4.4. Sanger Sequencing
4.5. ALDEFLUOR Assay
4.6. Colony Forming Assay
4.7. Apoptosis Assay
4.8. Live/Dead Assay
4.9. Reactive Oxygen Species
4.10. Alamar Blue Viability Assay
4.11. Lentiviral Transduction
4.12. Statistical Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Ajayi, G.O.; Ma, A.; Modarai, S.R.; Opdenaker, L.M.; Sims-Mourtada, J. CRISPR/Cas9 Targeting of Aldehyde Dehydrogenase 1A1 Reveals Heterogeneous Roles in Radiation Response and Redox Stress Across Clonal Lines in Triple-Negative Breast Cancer. Int. J. Mol. Sci. 2025, 26, 2303. https://doi.org/10.3390/ijms26052303
Ajayi GO, Ma A, Modarai SR, Opdenaker LM, Sims-Mourtada J. CRISPR/Cas9 Targeting of Aldehyde Dehydrogenase 1A1 Reveals Heterogeneous Roles in Radiation Response and Redox Stress Across Clonal Lines in Triple-Negative Breast Cancer. International Journal of Molecular Sciences. 2025; 26(5):2303. https://doi.org/10.3390/ijms26052303
Chicago/Turabian StyleAjayi, Grace O., Aihui Ma, Shirin R. Modarai, Lynn M. Opdenaker, and Jennifer Sims-Mourtada. 2025. "CRISPR/Cas9 Targeting of Aldehyde Dehydrogenase 1A1 Reveals Heterogeneous Roles in Radiation Response and Redox Stress Across Clonal Lines in Triple-Negative Breast Cancer" International Journal of Molecular Sciences 26, no. 5: 2303. https://doi.org/10.3390/ijms26052303
APA StyleAjayi, G. O., Ma, A., Modarai, S. R., Opdenaker, L. M., & Sims-Mourtada, J. (2025). CRISPR/Cas9 Targeting of Aldehyde Dehydrogenase 1A1 Reveals Heterogeneous Roles in Radiation Response and Redox Stress Across Clonal Lines in Triple-Negative Breast Cancer. International Journal of Molecular Sciences, 26(5), 2303. https://doi.org/10.3390/ijms26052303